The present invention relates to the fields of organic chemistry, biochemistry, medicinal chemistry, microbiology and medicine. In particular, it relates to organic compounds that are fungal efflux pump inhibitors.
The information provided and the references cited herein are not admitted, nor should they be construed, to be prior art to the present invention; rather, they are provided solely to assist the reader in understanding the present invention.
Fungal infections are relatively rare in immuno-competent patients. In fact, a number of Candida species are often present as benign commensal organisms in the digestive system of healthy individuals (Shepherd, et al., Ann. Rev. Microbiol., 1985, 39:579-614). Fungal infections, however, can be life threatening for immuno-compromised patients. There are three major groups of immuno-compromised individuals that are at risk: (1) cancer patients undergoing chemotherapy, (2) organ transplant patients being treated with immuno-suppressants, and (3) AIDS patients. Data from the National Nosocomial Infections Surveillance System conducted in the United States showed a 487 percent increase in Candida bloodstream infections between 1980 and 1989 (Rinaldi, et al., Antimicrob. Ag. Chemother., 1995, 39:1-8). Oropharyngeal candidiasis is the most common fungal infection complication associated with AIDS with up to 90% of AIDS patients having had at least one episode of the infection (Powderly, AIDS research and Human Retroviruses, 1994, 10:925-929).
There are relatively few clinically useful anti-fungal agents. Among those available are amphotericin B, flucytosine, fluconazole, itraconazole and ketoconazole (Odds, J. Antimicrob. Chemother., 1993, 31: 463-471). However, resistance to all of these drugs is developing rapidly. Take, for example, fluconazole.
Fluconazole is currently the most extensively used anti-fungal agent for the treatment of patients with severe candidiasis. It has higher water solubility and a longer plasma half-life than other azole fungicides and has relatively low toxicity. Between 1988 and 1993, fluconazole was used to treat over 15 million patients, including at least 250,000 AIDS patients (Hitchcock, Biochem. Soc. Trans., 1993, 21:1039-1047). Given such wide-spread use, it comes as no surprise that fluconazole-resistant Candida strains have been reported (Rex, et al., Antimicrob. Ag. Chemother., 1995, 39:1-8; Vanden Bossche, et al., 1994, supra). In some cases the resistance was found to be due to mutations in C. albicans itself while in other cases C. albicans was simply displaced by Candida species less susceptible to fluconazole, namely, C. glabrata and C. krusei (Odds, 1993, supra).
The mechanism of resistance to fluconazole appears to be multifaceted. In one study, amplification of the CYP51 gene (encoding the fluconazole target P-450 protein C14 demethylase) was implicated (Vanden Bossche, et al., Antimicrob. Agents and Chemother., 1994, 36: 2602-2610). In another study, resistance was correlated with the appearance of an altered P-450 target protein with decreased affinity for fluconazole (Hitchcock, Biochem Soc. Trans., 1993, 21:1039-1047). However, fluconazole resistance appears to be primarily due to decreased accumulation of the drug in resistant cells (Vanden Bossche, et al., 1994; Odds, 1993, supra). Species intrinsically resistant to fluconazole such as C. glabrata, C. krusei and Aspergillus fumigatus have also been shown to accumulate less fluconazole (Vanden Bossche, et al., 1994, supra). C. glabrata and C. krusei, on the other hand, have been shown to accumulate itraconazole and to be susceptible to that compound (Marichal et al., Mycoses, 1995, 38:111-117). Thus, it appears that both intrinsic and acquired resistance may be due to decreased drug accumulation in the cell. There are several ways in which a cell can manipulate the intracellular concentration of a compound. One is preventing the compound from gaining access to the interior of the cell in the first place. Another is metabolic decomposition of the compound once it is in the cell. A further means is simply excreting the intact compound before it can have any effect on the cell. This latter approach is called efflux and the cell components involved in efflux, i.e., membrane transporter proteins, are called efflux pumps.
Efflux pumps are ubiquitous in all types of cells, from bacterial to mammalian (Higgins, Ann. Rev. Cell Biol., 1992, 8:67-113). Efflux is driven either by the energy of ATP hydrolysis (ABC-transporter superfamily) or by proton transfer (Major Facilitator superfamily). Efflux pumps exhibit differing degrees of specificity.
Some efflux pumps are extremely specific, such as the TetA pump in gram-negative bacteria, which effluxes tetracycline only. Others are less specific; e.g., the MsrA protein in Staphyloccus aureus effluxes not only erythromycin but related macrolides as well. There are also efflux pumps that are quite general in their efflux capability, excreting a variety of structurally unrelated compounds from a cell. Many efflux pumps are clinically significant.
Resistance to chemotherapeutics in some mammalian cancer cells has been attributed to a multi-drug resistant efflux pump known as P-glycoprotein (Pgp) (Gottesman, et al., Ann. Rev. Biochem., 1993, 62:385-427). Pseudomonas aeruginosa, which causes respiratory infections, adventitious infection in burn patients, etc., uses Mex efflux pumps to eliminate quinolones, as well as other structurally unrelated antibiotics (Nikaido, Science, 1994, 264:382-388). Multiple-drug resistant (MDR) efflux pumps have been implicated in fluconazole resistance in C. albicans and C. glabrata (Parkinson, et al., Antimicrob. Agents Chemother., 1995, 39:1696-1699; Sanglard, et al., Antimicrob. Agents Chemother., 1995, 39:2378-2386; Albertson, et al., Antimicrob. Agents Chemother., 1996, 40:2835-2841).
Based on the above, it would clearly be desirable to be able to inhibit the activity of fungal efflux pumps so that anti-fungal agents can accumulate in fungal cells in sufficient quantity to exert their effect. The present invention provides compounds that achieve this goal.
The present invention relates to compounds that are fungal efflux pump inhibitors. When administered to a patient suffering from an infection caused by a fungal species that employs efflux pump(s) as a resistance mechanism, the compounds inhibit the activity of the pump(s) allowing a co-administrated anti-fungal agent to accumulate in sufficient concentration to inhibit fungal cells and treat the infection.
Thus, in one aspect the present invention relates to a compound having the chemical formula: 
or a pharmaceutically acceptable salt thereof, wherein:
A1 is carbon or nitrogen, provided that when A1 is nitrogen, R5 does not exist;
R2 and R3 are independently selected from the group consisting of hydrogen, halo and xe2x80x94O(1C-4C)alkyl;
R4 and R6 are independently selected from the group consisting of hydrogen, halo, xe2x80x94O(1C-4C)alkyl, xe2x80x94OCF3, and Oxe2x80x94CH2(3C-6C)cycloalkyl;
R5 is selected from the group consisting of hydrogen and 
xe2x80x83R7 and R8 are independently selected from the group consisting of hydrogen, halo, xe2x80x94Cxe2x89xa1N, xe2x80x94O(1C-4C)alkyl, xe2x80x94OCHF2, xe2x80x94OCF3 and, taken together, xe2x80x94OCH2Oxe2x80x94;
R1 is selected from the group consisting of xe2x80x94(1C-4C)alkyl, -(3C-6C)cycloalkyl, 
xe2x80x83wherein:
A4 is selected from the group consisting of xe2x80x94NH, oxygen and sulfur;
A2, A3 and A5 are independently selected from the group consisting of carbon and nitrogen provided that no more than two of A2, A3 and A5 are nitrogen at the same time;
or,
R1 is xe2x80x94C(O)(CH2)n(R22)R9, wherein,
n is 0, 1, 2 or 3;
R9 is selected from the group consisting of hydrogen, xe2x80x94OH, xe2x80x94(1C-4C)alkyl, -(3C-6C)cycloalkyl, xe2x80x94CH2(3C-6C)cycloalkyl, 
wherein:
A6, A7 and A8 are independently selected from the group consisting of carbon, oxygen, sulfur and NR15;
A9, A10 and A11 are independently selected from the group consisting of carbon and nitrogen;
R10 and R11are independently selected from the group consisting of hydrogen, -(1C-4C)alkyl, xe2x80x94SO2R16, xe2x80x94C(O)R16 and xe2x80x94C(O)OR16, wherein:
R16 is selected from the group consisting of hydrogen and -(1C-4C)alkyl wherein the alkyl group may be substituted with 1, 2, 3, or 4 fluorines;
R12 is selected from the group consisting of hydrogen, -(1C-4C)alkyl, -(3C-6C)cycloalkyl, xe2x80x94CH2(3C-6C)cycloalkyl, xe2x80x94C(O)O-(1C-4C)alkyl, xe2x80x94SO2R17 and xe2x80x94SO2NR18R19, wherein,
R17, R18 and R19 are independently selected from the group consisting of hydrogen and -(1C-4C)alkyl;
R14 and R15 are independently selected from the group consisting of hydrogen, -(1C-4C)alky and xe2x80x94NR10R11;
R22 is selected from the group consisting of hydrogen and (1C-4C)alkyl;
or,
R9 is xe2x80x94C(R16)(R20)(CH2)pNR10OR11, wherein:
p is 0, 1 or 2;
R20 is selected from the group consisting of hydrogen and -(1C-4C)alkyl, the alkyl group being optionally substituted with an entity selected from the group consisting of xe2x80x94OH, xe2x80x94O(1C-4C)alkyl, xe2x80x94Cxe2x89xa1N, xe2x80x94SO2(1C-4C)alkyl and 
and, the compound comprises a racemic mixture, a pure enantiomer or a pure atropisomer of either the racemic mixture or the pure enantiomer.
In an aspect of this invention, the compound is in the S absolute configuration at the starred carbon.
In an aspect of this invention, in the above compounds, A1 is carbon and R22 is selected from the group consisting of hydrogen and xe2x80x94CH3.
In an aspect of this invention, in the above compounds, A1 is nitrogen.
In an aspect of this invention, in the above compounds, R4 and R6 are independently selected from the group consisting of xe2x80x94O(1C-4Calkyl) and xe2x80x94OCH2(3C-6C)cycloalkyl.
In an aspect of this invention, in the above compounds, R4 and R6 are OCH3.
In an aspect of this invention, in the above compounds, R7 is selected from the group consisting of hydrogen and halogen and R8 is hydrogen.
In an aspect of this invention, in the above compounds, R7 is fluorine.
In an aspect of this invention, in the above compounds, R6 is selected from the group consisting of xe2x80x94OCH3 and 
and,
R7 is F.
In an aspect of this invention, in the above compounds, R6 is selected from the group consisting of xe2x80x94OCH3 and 
and,
R5 is xe2x80x94C(O)CH3.
In an aspect of this invention, in the above compounds, R4 and R6 are 
In an aspect of this invention, in the above compounds, R21 is xe2x80x94NHSO2CH3.
In an aspect of this invention, in the above compounds, R5 is 
An aspect of this invention is a method for inhibiting a fungal cell that employs an efflux pump resistance mechanism, comprising contacting the cell with an anti-fungal agent and any one of the above compounds.
In an aspect of this invention, in the above method, the anti-fungal agent is an azole anti-fungal agent.
In an aspect of this invention, in the above method, the azole fungicide is selected from the group consisting of fluconazole and posaconazole.
In an aspect of this invention, in the above method, the fungal cell is first contacted with the compound and then with the anti-fungal agent.
In an aspect of this invention, in the above method, the fungal cell is contacted with the compound and the anti-fungal agent simultaneously.
In an aspect of this invention, in the above method, the fungal cell is a genus Candida cell.
In an aspect of this invention, in the above method, the fungal cell is a genus Aspergillus cell.
An aspect of this invention is a method for treating an infection caused by a fungus that employs an efflux pump resistance mechanism, comprising administering to a patient in need thereof a therapeutically effective amount of an anti-fungal agent and an one of the above compounds.
In an aspect of this invention, in the above method, the infection is caused by a genus Candida fungus.
In an aspect of this invention, in the above methods, the Candida fungus is C albicans, C. krusei, C. tropicalis, C. parapsilosis or C. glabrata. 
In an aspect of this invention, in the above methods, the infection is caused by a genus Aspergillus fungus.
In an aspect of this invention, in the above methods, the genus Aspergillus fungus is Aspergillus fumigatus. 
In an aspect of this invention, in the above method, the compound and the anti-fungal agent are contacted with the fungal cell or administered to the patient simultaneously.
In an aspect of this invention, in the above method, the compound is contacted with the fungal cell or administered to the patient first followed by contact with or administration of the anti-fungal agent.
An aspect of this invention is a pharmaceutical composition comprising a pharmaceutically acceptable carrier or excipient and any of the above compounds.
In an aspect of this invention, the above pharmaceutical composition further comprises a therapeutically effective amount of an anti-fungal agent.
In an aspect of this invention, in the above pharmaceutical composition, the anti-fungal agent is an azole anti-fungal agent.
In an aspect of this invention, in the above pharmaceutical composition, the azole anti-fungal agent is fluconazole or posaconazole.